The Hepatitis B virus (HBV) remains a major cause of chronic hepatitis and hepatocellular carcinoma (HCC) globally. HBV is broadly classified into ten genotypes (A-J). Each genotype is distinctive due to its geographical distribution, response to certain treatments, and association with disease severity. The investigation of HBV genotypes is worthwhile as it will help predict patient prognosis and ensure effective treatment. Therefore, this study aimed to determine the prevalent HBV genotypes in the multiethnic population of Karachi, Pakistan. A total of 206 patients were enrolled in the study from January 2011 to April 2016. Of the 206 patients that we investigated, a total of 159 patients were positive for HBV infection. Furthermore, genotyping analysis of the positive samples revealed that the most common genotypes were HBV D (59% samples (n=94)), co-infection with genotype A & D (22% (n=35)) and genotype A (16% (n=26)). Most of the HBV-positive patients were males between 21 to 40 years of age. As genotype D is associated with worse outcomes, it is imperative that HBV awareness programs are launched to prevent the further spread of infections in this densely populated city.